Dr. Bilgen is a current CDA-1 recipient (2008) and has a strong bioengineering background with hands-on research experience over 10 years in cartilage tissue engineering. Dr. Bilgen's research interests include bioengineering applications to clinical problems in orthopaedics, specifically skeletal biology, repair and regeneration. This application aims to develop a transplantable biocomposite to restore cartilage of a traumatized joint. Limb trauma in young veterans often results in loss of cartilage tissue function that leads to severe arthritis and osteoarthritis. Total national cost of arthritis was $81 billion in medical costs and $47 billion in lost wages in 2003. US veterans face increased risks of arthritis compared with non-veterans due to orthopedic injuries at the military. Implantation of engineered cartilage that is produced by the patient's autologous chondrocytes is not fully successful in healing cartilage defects. This project addresses two main problems in cartilage tissue engineering: 1) limited availability of adult autologous chondrocytes and limited differentiation of mesenchymal stem cells (MSC), 2) insufficient mechanical properties of engineered cartilage. Due to limited chondrocyte availability, adult mesenchymal stem cells (MSC) are widely used to produce cartilage tissue, but they do not secrete cartilage matrix at the same rate as chondrocytes. We and others have shown that co-culturing adult stem cells with chondrocytes improve the biochemical composition of engineered cartilage. Application of intermittent mechanical loading such as compression or shear during tissue cultivation has been shown to improve cartilage formation, however the optimum dose differs with cell and tissue properties. The central hypothesis is that the combination of co-culturing chondrocytes and MSC and application of biaxial mechanical loading will synergistically enhance deposition of extracellular matrix (ECM) and improve the functional properties of cartilage tissue after transplantation. Over the five-year CDA-2 program, Dr. Bilgen aims to achieve three research goals: 1- Develop a co-culture system to address the limited availability of chondrocytes: The approach is to optimize ECM deposition using a combination of chondrocytes and synovial MSC and growth factors. The expected outcome is a novel cell sourcing methodology that utilizes co-cultures of chondrocytes and synovial MSC. 2- Create a cartilage biocomposite with improved mechanical properties: We designed a novel biaxial loading device, which was shown to be compatible with the development of tissue-engineered cartilage biocomposites in long term in vitro studies. The approach is to investigate the effects of uniaxial and biaxial mechanical loading in vitro on the differentiation of cells and the mechanical properties of engineered tissue. The expected outcome is the development of a novel biaxial loading system to create a biocomposite that can successfully repair and restore function to cartilage. 3- Develop an in vivo-compatible biocomposite for cartilage repair: The approach will be to cultivate the biocomposite in vitro to improve functional properties followed by implantation in vivo in a chondral defect in a minipig model. The expected outcome is the development of a biocomposite that can successfully repair cartilage in vivo. The innovation in this grant proposal lies in the synergistic application of instructive co- culturing and a novel mechanical loading scheme on a cartilage tissue engineering system using mesenchymal stem cells. The broader impact is that the developed techniques are anticipated to treat degenerative joint diseases and prevent joint replacement surgeries, thus impacting nearly half of the population.